Recombinant fungal strains with reduced levels of mycotoxins

Abstract

Described herein are recombinant edible filamentous fungal strain comprising a genetic modification of a gene in a mycotoxin biosynthesis pathway and methods to produce such recombinant edible filamentous fungal strains. Such recombinant strains comprise reduced levels of mycotoxins. Also described are food materials comprising the recombinant edible filamentous fungal strains described herein, and methods to prepare them.

1 . A recombinant filamentous fungal strain, wherein the recombinant strain comprises a genetic modification of a gene in a mycotoxin biosynthesis pathway and wherein the recombinant strain produces a reduced level of the mycotoxin. 
     
     
         2 . The recombinant filamentous fungal strain of  claim 1 , wherein the mycotoxin is a polyketide. 
     
     
         3 . The recombinant filamentous fungal strain of  claim 1 , wherein the mycotoxin is selected from the group consisting of a fumonisin, a fusarin, fusaric acid and a combination thereof. 
     
     
         4 . The recombinant filamentous fungal strain of  claim 1 , wherein the recombinant strain comprises a modification of a gene encoding a polyketide synthase. 
     
     
         5 . The recombinant filamentous fungal strain of  claim 4 , wherein the gene encoding a polyketide synthase is selected from the group consisting of Fum1, Fus1, Fub1 and a combination thereof. 
     
     
         6 . The recombinant filamentous fungal strain of  claim 5 , wherein the recombinant strain comprises a modification of the Fum1 gene and the recombinant strain produces a reduced level of fumonisin; or the recombinant strain comprises a modification of the Fus1 gene and the recombinant strain produces a reduced level of Fusarin C; or the recombinant strain comprises a modification of the Fub1 gene and the recombinant strain produces a reduced level of Fusaric acid; or a combination thereof. 
     
     
         7 . The recombinant filamentous fungal strain of  claim 1 ,
 wherein the mycotoxin is fumonisin and the genetic modification comprises a modification of one or more of the genes selected from the group consisting of Fum1, Fum2, Fum3, Fum6, Fum7, Fum8, Fum10, Fum11, Fum13, Fum14, Fum15, Fum16, Fum17, Fum18, Fum19, Fum20 and Fum21; or   wherein the mycotoxin is fusarin and the genetic modification comprises a modification of one or more of the genes selected from the group consisting of Fus1, Fus2, Fus3, Fus4, Fus5, Fus6, Fus7, Fus8 and Fus9; or   wherein the mycotoxin is fusaric acid and the genetic modification comprises a modification of one or more of the genes selected from the group consisting of Fub1, Fub2, Fub3, Fub4, Fub5, Fub6, Fub7, Fub8, Fub9, Fub10, Fub 11 and Fub12;   or a combination thereof.   
     
     
         8 . (canceled) 
     
     
         9 . The recombinant filamentous fungal strain of  claim 1 , wherein the genetic modification of the gene comprises a partial or complete deletion of the gene. 
     
     
         10 . The recombinant filamentous fungal strain of  claim 1 , wherein the genetic modification of the gene is performed using at least one of homologous recombination, CRISPR, and TALEN. 
     
     
         11 - 12 . (canceled) 
     
     
         13 . The recombinant filamentous fungal strain of  claim 1 , wherein the genetic modification of the gene is performed in combination with introducing a selectable marker. 
     
     
         14 . The recombinant filamentous fungal strain of  claim 13 , wherein the selectable marker is hygromycin resistance. 
     
     
         15 . (canceled) 
     
     
         16 . The recombinant filamentous fungal strain of  claim 1 , wherein the genetic modification comprises a modification of the Fum1, Fum6 or Fum8 gene. 
     
     
         17 . The recombinant filamentous fungal strain of  claim 16 , wherein the genetic modification comprises a modification of the Fum1 gene. 
     
     
         18 . The recombinant filamentous fungal strain of  claim 17 , wherein the Fum1 gene is at least 70% identical to SEQ ID NO: 2. 
     
     
         19 . (canceled) 
     
     
         20 . The recombinant filamentous fungal strain of  claim 17 , wherein the Fum1 comprises SEQ ID NO: 2. 
     
     
         21 . The recombinant filamentous fungal strain of  claim 1 , wherein the filamentous fungus is a  Fusarium  species. 
     
     
         22 . The recombinant filamentous fungal strain of  claim 21 , wherein the filamentous fungus is selected from the group consisting of MK7 (ATCC PTA-10698) and  Fusarium venenatum.    
     
     
         23 . (Canceled) 
     
     
         24 . The recombinant filamentous fungal strain of  claim 1 , wherein the recombinant strain comprises less than about 10 ppm of a mycotoxin selected from the group consisting of fumonisin, fusarin or fusaric acid and a combination thereof on a dry weight basis. 
     
     
         25 - 26 . (canceled) 
     
     
         27 . The recombinant edible-filamentous fungal strain of  claim 1 , wherein the fumonisin comprises funonisin FB. 
     
     
         28 . (canceled) 
     
     
         29 . The recombinant filamentous fungal strain of  claim 1 , wherein the fusarin comprises fusarin C. 
     
     
         30 . A biomat comprising the recombinant filamentous fungal strain of  claim 1 . 
     
     
         31 . A method of producing the bioniat of  claim 30  comprising
 (a) inoculating an artificial liquid growth medium with an effective amount of the recombinant filamentous fungal strain; 
 (b) incubating the inoculated filamentous fungi to produce a filamentous fungal biomat on the surface of the liquid medium; and 
 (c) harvesting the filamentous fungal biomat. 
 
     
     
         32 - 34 . (canceled)